Breast cancer afflicts one in eight women in the United States over their lifetime (1). ERα (NR3A1, (2)) mediates estrogen responsiveness (reviewed in (3)) and plays crucial roles in the etiology of breast cancer (reviewed in (4)). It has been developed into the single most important genetic biomarker and target for breast cancer therapy. ERα is present at detectable levels by ligand-binding and immunohistochemical assays in approximately 75% of clinical breast cancers. Selection of patients with ERα-positive breast tumors increases endocrine-based therapy response rates from about one-third in unselected patients to about one-half in patients with ERα-positive tumors (5). Since expression of PgR is dependent upon ERα activity, further selection of patients with ERα- and PgR-positive tumors enhances the breast cancer hormonal therapy response rate to nearly 80% (5). Although ERβ (NR3A2 (2)) also mediates responses to estrogens (reviewed in (3)), its roles in breast cancer are not as well understood. Reports have linked ERβ expression with low tumor aggressiveness (6) and higher levels of proliferation markers in the absence of ERα (7).
Members of the ErbB family of transmembrane tyrosine kinase receptors have been implicated in the pathogenesis of breast cancer. The members include EGFR (also HER1; ErbB1), ErbB2 (HER2; Neu), ErbB3 (HER3) and ErbB4 (HER4) (reviewed in (8)). ErbB members stimulate signal transduction pathways that involve MAPK. In response to initial binding of EGF-like peptide hormones, ErbB members form homodimers and heterodimers in various combinations to recruit distinct effector proteins (reviewed in (9)). Although ErbB2 has not been demonstrated to interact directly with peptide hormones, it serves as a common regulatory heterodimer subunit with other ligand-bound ErbB members (reviewed in (10, 11)). Unlike the other ErbB members, ErbB3 lacks intrinsic kinase activity and, therefore, is required to heterodimerize with other ErbB members to participate in signaling (12).
Independent overexpression of either EGFR (reviewed in (13)) or ErbB2 (reviewed in (14)) associates with ER-negative tumor status, indicates aggressive tumor behavior, and predicts poor prognosis. Moreover, patients whose tumors coexpress both EGFR and ErbB2 exhibit a worse outcome than patients with tumors that overexpress only one of these genes (15, 16). Overexpression of ErbB2, most often due to gene amplification, occurs in approximately 15-30% of all breast cancers ((17), reviewed in (14)). Some (18-23), but not all reports (24, 25), have implicated ErbB2 in the development of resistance to antiestrogens.
ErbB2 has been targeted for development of the successful clinical agent Herceptin (trastuzumab), a recombinant humanized monoclonal antibody directed against this receptor's ectodomain (reviewed in (26)). Herceptin has been shown to be a suitable option as a first-line single-agent therapy (27), but will likely prove most beneficial as an adjuvant (28, 29). Clinical trials are currently underway to evaluate the combination of Herceptin with antiestrogens as a rational approach to treating ERα-positive/ErbB2-overexpressing tumors (23). In the near future, Herceptin will also likely be evaluated in combination with the small molecule EGFR tyrosine kinase inhibitor ZD 1829 (Iressa), since this ATP-mimetic has been shown to almost completely block transphosphorylation of ErbB2 via heterodimerization with EGFR in intact cells (30) and inhibits the growth of breast cancer cell lines overexpressing both EGFR and ErbB2 (31). Hence, a combination of ZD1829 and Herceptin may be particularly beneficial to those patients whose tumors coexpress EGFR and ErbB2.
The ability of ErbB3 and ErbB4 to predict clinical course is not as clearly recognized as that of EGFR and ErbB2. ErbB3 has been observed at higher levels in breast tumors than normal tissues, showing associations with unfavorable prognostic indicators including ErbB2 expression (32), lymph node-positive status (33), and tumor size. However, it also associated with ERα-positive status, a favorable marker of hormonal sensitivity (34). In contrast, ErbB4 has shown associations with only positive indicators. ErbB4 overexpression has been associated with ERα-positive status (34, 35), more differentiated histotypes (36), a more favorable outcome (16) and it may oppose the negative effects of ErbB2 on clinical course (16).
Despite the utility of ERs and ErbB members as indicators of clinical course, there remains a great need to identify additional breast cancer biomarkers. A family of potential candidate biomarkers includes the orphan nuclear receptors ERRα (37-39), ERRβ (37, 40), and ERRγ (40-42) (NR3B1, NR3B2, and NR3B3, respectively (2)). These receptors share significant amino acid sequence identity with ERα and ERβ. They also exhibit similar but distinct biochemical and transcriptional activities as the ERs. Each of the ERRs has been demonstrated to bind and activate transcription via consensus palindromic EREs (43-46) in addition to ERREs (39, 42, 44, 47-50), which are composed of an ERE half-site with a 5′ extension of 3 base pairs. However, whereas ERs are ligand-activated transcription factors, the ERRs do not bind natural estrogens (37, 51). Instead, the ERRs may serve as constitutive regulators, interacting with transcriptional coactivators in vitro in the absence of ligands (45, 50, 52). Bulky amino acid side chains in the ligand binding pockets of ERRs substitute for the analogous ligand-induced interactions observed in ERα (52, 53). However, the ligand-binding pockets of the ERRs still allow binding of the synthetic estrogen diethylstilbestrol, but as an antagonist because it also disrupts coactivator interactions with ERRs (51). Similarly, the selective estrogen receptor modulator (SERM) 4-hydroxytamoxifen selectively antagonizes ERRγ in cell-based assays (46, 52, 54).
The transcriptional activity of each ERR depends upon the promoter and the particular cell line in which it is assayed as well as the presence of ERs. (42-46, 48-51, 53-62) For example, whereas ERRα stimulates ERE-dependent transcription in the absence of ERα in HeLa cells, it down-modulates E2-stimulated transcription in ERα-positive human mammary carcinoma MCF-7 cells via an active mechanism of repression. (43) ERRα can also modulate transcription of at least some genes that are estrogen responsive and/or implicated in breast cancer such as pS2 (55), aromatase (59), osteopontin (57, 58) and lactoferrin (56, 61). Thus, the ERRs likely play important roles in at least some breast cancers by modulating or substituting for ER-dependent activities.